The present invention relates generally to taxane-oligomer conjugates and to methods for making and using such conjugates. The taxane-oligomer conjugates of the invention operate as prodrugs, hydrolyzing under normal physiological conditions to provide therapeutically active taxanes, such as paclitaxel or docetaxel. The taxane-oligomer conjugates exhibit improved solubility characteristics, improved oral bioavailability, and an improved pharmacokinetic profile. The present invention also relates to pharmaceutical compositions comprising these taxane-oligomer conjugates and to methods of making and using such taxane-oligomer conjugates and pharmaceutical compositions.
Paclitaxel (Taxol) is a natural diterpene product isolated from the pacific yew tree (Taxus brevifolia). Wani et al. first isolated paclitaxel in 1971 by chemical and X-ray crystallographic methods. Paclitaxel is a complex diterpene having a taxane ring with a 4-membered oxetane ring and an ester side chain at position C-13. The complex structure of paclitaxel is as follows: 
Paclitaxel has been approved for clinical use in the treatment of refractory ovarian cancer in the United States. (Markman 1991; McGuire et al. 1989). Paclitaxel has also been approved for treatment of breast cancer. (Holmes et al. 1991) Additionally, paclitaxel is a candidate for treatment of neoplasms of the skin (Einzig et al.) and head and neck carcinomas (Forastire et al. 1990). Paclitaxel is also useful for the treatment of polycystic kidney disease (Woo et al. 1994), lung cancer and malaria.
Paclitaxel mediates its anti-cancer effects by lowering the critical concentration of tubulin necessary for microtubule formation. Microtubules are polymers of tubulin in dynamic equilibrium with tubulin heterodimers that are composed of xcex1 and xcex2 protein subunits. Paclitaxel shifts the equilibrium towards microtubule assembly. Paclitaxel-induced microtubules are excessively stable, thereby inhibiting dynamic reorganization of the microtubule network, and resulting in microtubule bundles that form during all phases of the cell cycle and numerous abnormal mitotic asters that are not associated with centrioles.
Paclitaxel entered Phase I clinical trials in 1983, but immediately encountered formulation difficulties due to its aqueous insolubility. This difficulty was partially overcome by formulating Paclitaxel as an emulsion with Cremophor EL(copyright). However, since paclitaxel must be given at relatively high dosages, large amounts of Cremophor EL(copyright) are required. When administered intravenously, such formulations can produce vasodilatation, labored breathing, lethargy, hypertension and death in dogs, and are also believed to be responsible for the allergic-type reactions observed during paclitaxel administration in humans. Accordingly, there is a need in the art for a means for administering paclitaxel which increases its water solubility and thereby avoids the need for formulating paclitaxel with potentially allergenic emulsion reagents.
Efforts to overcome the allergy problems of formulated paclitaxel have thus far been directed at lengthening the infusion time and premedicating patents with immunosuppressive agents, such as glucocorticoids and also with antihistamines. These agents have their own set of side effects and are an added cost to the already expensive cost of cancer treatment. Furthermore, while such agents have been shown to reduce the incidence and severity of hypersensitivity reactions, they are not fully protective. (Rowinsky et al. 1992). Accordingly, there is a need in the art for means for administering paclitaxel which avoids lengthened infusion times and the allergic reactions associated with emulsion reagents and thereby also avoids the need for such adjunctive treatment.
Several groups have investigated the synthesis of prodrug forms of paclitaxel. (Taylor 1994); (Kingston, D. G. 1991). Prodrugs are inactive or partially inactive chemical derivatives of drugs that are metabolized to yield the pharmacologically active drug. Studies have been directed toward synthesizing paclitaxel analogs where the 2xe2x80x2 and/or 7-position is derivatized with groups that enhance water solubility. These efforts have yielded prodrug compounds that are more water-soluble than the parent compound while displaying the cytotoxic properties of paclitaxel upon activation. For example, increased water-solubility has been achieved by derivatizing paclitaxel with high molecular weight polyethylene glycol (PEG) polymers. (See Greenwald, et al. 1996; Greenwald et al. 1995). However, while these derivatized paclitaxel compounds have increased solubility, they also result in a corresponding decrease in drug load, due to the high molecular weight PEG necessary to achieve adequate solubility. Accordingly, there is a need in the art for taxane prodrugs which improve paclitaxel solubility without drastically increasing the molecular weight of the paclitaxel compound.
Efficient utilization of prodrugs, especially taxane prodrugs, requires that the properties of the prodrug must be adequately balanced to achieve a useful pharmacokinetic profile. In one aspect, it is desirable for the prodrug to be hydrophilic in order to enhance the ability to formulate the prodrug. On the other hand, the prodrug must be appropriately hydrophobic to permit interaction of the prodrug with biological membranes. There is therefore a need in the art for taxane prodrugs that accommodate the foregoing disparate requirements for useful therapeutic agents.
The present inventors have surprisingly and unexpectedly discovered taxane-oligomer compounds and salts of such compounds (collectively referred to herein as xe2x80x9ctaxane prodrugsxe2x80x9d) that significantly increase the water-solubility of taxane drugs without drastically increasing their molecular weight. The taxane prodrugs described herein eliminate the need for microemulsion formulation using Cremophor EL(copyright).
Embodiments according to the present invention provide a taxane prodrug comprising a taxane joined by a hydrolyzable bond to an oligomer having the following formula: 
wherein n is from 1 to 12, m is from 1 to 25, p is from 2 to 12, X+ is a positive ion and Zxe2x88x92 is a negative ion.
In other embodiments of the present invention, the taxane is paclitaxel or a paclitaxel analog which retains some or all of the therapeutic activity of paclitaxel. Taxane may also be docetaxel.
The taxane prodrug may be derivatized by as many oligomers as there are sites on the taxane for attachment of such oligomers. For example, paclitaxel has 3 attachment sites (hydroxyl groups) and can therefore be derivatized by 1, 2 or 3 of the oligomers. Similarly, docetaxel paclitaxel has 4 attachment sites (hydroxyl groups) and can therefore be derivatized by 1, 2, 3 or 4 of the oligomers.
In another aspect, the taxane prodrugs can be delivered via oral administration to provide a therapeutically effective dose of the taxane to the bloodstream. Furthermore, the orally delivered prodrugs can provide a therapeutically effective dose of the taxane to the target organ or tissue.
Embodiments of the present invention also provide pharmaceutical compositions comprising the taxane prodrugs of the invention in association with a pharmaceutically acceptable carrier. Such pharmaceutical compositions may be formulated so as to be suitable for oral administration, and may be in a dosage form selected from the group consisting of: tablets, capsules, caplets, gelcaps, pills, liquid solutions, suspensions or elixirs, powders, lozenges, micronized particles and osmotic delivery systems. Methods for treating a mammalian subject having a paclitaxel-responsive disease condition are also provided. The mammalian subject is preferably a human.
In one aspect of the methods of treatment, the taxane prodrug is delivered via an oral route of administration to provide a therapeutically effective dose of the taxane into the bloodstream. In another aspect, the taxane prodrug is delivered via a parenteral route of administration, providing a therapeutically effective dose of the taxane to target organs and/or tissues. In yet another aspect, the taxane prodrug is delivered via an oral route of administration, providing a therapeutically effective dose of the taxane to target organs and/or tissues. Furthermore, the taxane prodrug may be administered in association with a pharmaceutically acceptable carrier.
In a further aspect, the taxane-responsive disease condition treated according to the therapeutic methods of the invention is selected from the group consisting of benign and malignant neoplasms, and may include hepatocellular carcinoma, urogenital carcinoma, liver metastases, gastrointestinal cancers, lymphoma, leukemia, melanoma, Kaposi""s sarcoma, and cancers of the pancreas, kidney, cervix, breast, ovary, brain, and prostate. In one aspect, the disease condition comprises ovarian cancer and the taxane prodrug is administered optionally with cisplatin, either simultaneously or sequentially. In another aspect, the disease condition comprises breast cancer and the taxane prodrug is administered optionally with doxorubicin, either simultaneously or sequentially.
As used herein the term xe2x80x9cPEGxe2x80x9d refers to straight or branched polyethylene glycol oligomer and monomers and also includes polyethylene glycol oligomers that have been modified to include groups which do not eliminate the amphiphilic properties of such oligomer, e.g. without limitation, alkyl, lower alkyl, aryl, amino-alkyl and amino-aryl. The term xe2x80x9cPEG subunitxe2x80x9d refers to a single polyethylene glycol unit, i.e., xe2x80x94(CH2CH2O)xe2x80x94.
As used herein, the term xe2x80x9clower alkylxe2x80x9d refers to a straight or branched chain hydrocarbon having from one to 8 carbon atoms.
As used herein, terms such as xe2x80x9cnon-hydrolyzablexe2x80x9d and phrases such as xe2x80x9cnot hydrolyzablexe2x80x9d are used to refer to bonds which cannot be hydrolyzed under normal physiological conditions, as well as bonds which are not rapidly hydrolyzed under normal physiological conditions such as carbamate and amide bonds. The term xe2x80x9chydrolyzablexe2x80x9d refers to bonds which are hydrolyzed under physiological conditions. In a preferred aspect of the invention, 50% of the taxane prodrug is hydrolyzed in a normal population within 4 hours after intravenous administration.
A xe2x80x9ctherapeutically effective amountxe2x80x9d is an amount necessary to prevent, delay or reduce the severity of the onset of disease, or an amount necessary to arrest or reduce the severity of an ongoing disease, and also includes an amount necessary to enhance normal physiological functioning.
As used herein, a xe2x80x9cpharmaceutically acceptablexe2x80x9d component (such as a salt, carrier, excipient or diluent) of a formulation according to the present invention is a component which (1) is compatible with the other ingredients of the formulation in that it can be combined with the taxane prodrugs of the present invention without eliminating the biological activity of the taxane prodrugs; and (2) is suitable for use with an animal (e.g., a human) without undue adverse side effects, such as toxicity, irritation, and allergic response. Side effects are xe2x80x9cunduexe2x80x9d when their risk outweighs the benefit provided by the pharmaceutical composition. Examples of pharmaceutically acceptable components include, without limitation, standard pharmaceutical carriers, such as phosphate buffered saline solutions, water, emulsions such as oil/water emulsions, microemulsions, and various types of wetting agents.
As used herein, the term xe2x80x9ctaxanexe2x80x9d is used to refer to a class of compounds having a basic three ring structure which includes rings A, B and C of paclitaxel: 
including, without limitation, paclitaxel and paclitaxel analogs which retain some or all of the anti-cancer activity of paclitaxel.
FIG. 1 shows the conversion of acetate (Compound 4) (see Examples below) to paclitaxel over the course of 26 days at room temperature, where the acetate (Compound 4) was dissolved in wet acetonitrile. FIG. 1 shows that acetate (Compound 4) smoothly converted quantitatively into paclitaxel with no significant side product formation, as observed by analytical HPLC analysis.
FIG. 2 shows in vitro hydrolysis of acetate (Compound 4) in heparinized rat plasma and demonstrates that most of the taxane prodrug rapidly hydrolyzes within two hours to provide free paclitaxel.
FIG. 3 shows percent growth of NIH:OVCAR-3 cells treated with compounds of the present invention in comparison with paclitaxel.
FIG. 4 shows caspase-3 activation by NIH:OVCAR-3 cells stimulated with compounds of the present invention in comparison with paclitaxel.